Showing papers on "Tartrate-resistant acid phosphatase published in 2018"

Suppression Effect of Astaxanthin on Osteoclast Formation In Vitro and Bone Loss In Vivo.

Abstract: Osteoporosis is characterized by a reduction of the bone mineral density (BMD) and microarchitectural deterioration of the bone, which lead to bone fragility and susceptibility to fracture. Astaxanthin (AST) has a variety of biological activities, such as a protective effect against asthma or neuroinflammation, antioxidant effect, and decrease of the osteoclast number in the right mandibles in the periodontitis model. Although treatment with AST is known to have an effect on inflammation, no studies on the effect of AST exposure on bone loss have been performed. Thus, in the present study, we examined the antiosteoporotic effect of AST on bone mass in ovariectomized (OVX) mice and its possible mechanism of action. The administration of AST (5, 10 mg/kg) for 6 weeks suppressed the enhancement of serum calcium, inorganic phosphorus, alkaline phosphatase, total cholesterol, and tartrate-resistant acid phosphatase (TRAP) activity. The bone mineral density (BMD) and bone microarchitecture of the trabecular bone in the tibia and femur were recovered by AST exposure. Moreover, in the in vitro experiment, we demonstrated that AST inhibits osteoclast formation through the expression of the nuclear factor of activated T cells (NFAT) c1, dendritic cell-specific transmembrane protein (DC-STAMP), TRAP, and cathepsin K without any cytotoxic effects on bone marrow-derived macrophages (BMMs). Therefore, we suggest that AST may have therapeutic potential for the treatment of postmenopausal osteoporosis.

The Lysosomal Protein Arylsulfatase B Is a Key Enzyme Involved in Skeletal Turnover

Abstract: Skeletal pathologies are frequently observed in lysosomal storage disorders, yet the relevance of specific lysosomal enzymes in bone remodeling cell types is poorly defined. Two lysosomal enzymes, ie, cathepsin K (Ctsk) and Acp5 (also known as tartrate-resistant acid phosphatase), have long been known as molecular marker proteins of differentiated osteoclasts. However, whereas the cysteine protease Ctsk is directly involved in the degradation of bone matrix proteins, the molecular function of Acp5 in osteoclasts is still unknown. Here we show that Acp5, in concert with Acp2 (lysosomal acid phosphatase), is required for dephosphorylation of the lysosomal mannose 6-phosphate targeting signal to promote the activity of specific lysosomal enzymes. Using an unbiased approach we identified the glycosaminoglycan-degrading enzyme arylsulfatase B (Arsb), mutated in mucopolysaccharidosis type VI (MPS-VI), as an osteoclast marker, whose activity depends on dephosphorylation by Acp2 and Acp5. Similar to Acp2/Acp5-/- mice, Arsb-deficient mice display lysosomal storage accumulation in osteoclasts, impaired osteoclast activity, and high trabecular bone mass. Of note, the most prominent lysosomal storage accumulation was observed in osteocytes from Arsb-deficient mice, yet this pathology did not impair production of sclerostin (Sost) and Fgf23. Because the influence of enzyme replacement therapy (ERT) on bone remodeling in MPS-VI is still unknown, we additionally treated Arsb-deficient mice by weekly injection of recombinant human ARSB from 12 to 24 weeks of age. We found that the high bone mass phenotype of Arsb-deficient mice and the underlying bone cell deficits were fully corrected by ERT in the trabecular compartment. Taken together, our results do not only show that the function of Acp5 in osteoclasts is linked to dephosphorylation and activation of lysosomal enzymes, they also provide an important proof-of-principle for the feasibility of ERT to correct bone cell pathologies in lysosomal storage disorders. © 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals Inc.

Attributes of Bio-Oss® and Moa-Bone® graft materials in a pilot study using the sheep maxillary sinus model.

Abstract: Background and objective The aim of this pilot study was to characterize surface morphology and to evaluate resorption and osseous healing of two deproteinated bovine bone graft materials after sinus grafting in a large animal model. Material and methods Surfaces of a novel particulate bovine bone graft, Moa-Bone® were compared with Bio-Oss® using scanning electron microscopy. Six sheep then had maxillary sinus grafting bilaterally, covered with BioGide® . Grafted maxillae were harvested after 4, 6 and 12 weeks. Healing was described for half of each site using resin-embedded ground sections. For the other half, paraffin-embedded sections were examined using tartrate resistant acid phosphatase staining for osteoclast activity, runt-related transcription factor2 immunohistochemistry for pre-osteoblasts and osteoblasts and proliferating cell nuclear antigen for proliferative cells. Results Moa-Bone® had a smoother, more porous fibrous structure with minimal globular particles compared with Bio-Oss® . After 4 weeks, woven bone formed on both grafts and the Moa-Bone® particles also showed signs of resorption. After 12 weeks, Moa-Bone® continued to be resorbed, however Bio-Oss® did not; both grafts were surrounded by maturing lamellar bone. Moa-Bone® was associated with earlier evidence of runt-related transcription factor 2-positive cells. Moa-Bone® but not Bio-Oss® was associated with strong tartrate resistant acid phosphatase-positive osteoclasts on the graft surface within resorption lacunae at both 4 and 6 weeks post-grafting. Conclusion Both materials supported osseous healing and maturation without inflammation. Moa-Bone® showed marked osteoclast activity after 4 and 6 weeks and demonstrated positive attributes for grafting, if complete remodeling of the graft within the site is desired. Further optimization of Moa-Bone® for maxillofacial applications is warranted.

Inhibitory effects of low intensity pulsed ultrasound on osteoclastogenesis induced in vitro by breast cancer cells.

Abstract: Bone tissue is one of the main sites for breast metastasis; patients diagnosed with advanced breast cancer mostly develop bone metastasis characterized by severe osteolytic lesions, which heavily influence their life quality. Low Intensity Pulsed Ultrasound (LIPUS) is a form of mechanical energy able to modulate various molecular pathways both in cancer and in health cells. The purpose of the present study was to evaluate for the first time, the ability of LIPUS to modulate osteolytic capability of breast cancer cells. Two different approaches were employed: a) Indirect method -conditioned medium obtained by MDA-MB-231 cell line treated or untreated with LIPUS was used to induce osteoclast differentiation of murine macrophage Raw264.7 cell line; and b) Direct method -MDA-MB-231 were co-cultured with Raw264.7 cells and treated or untreated with LIPUS. LIPUS treatment impaired MDA-MB-231 cell dependentosteoclast differentiation and produced a reduction of osteoclast markers such as Cathepsin K, Matrix Metalloproteinase 9 and Tartrate Resistant Acid Phosphatase, suggesting its role as an effective and safe adjuvant in bone metastasis management. LIPUS treatment could be a good and safety therapeutic adjuvant in osteolyitic bone metastasis not only for the induction properties of bone regeneration, but also for the reduction of osteolysis.

Tartrate-resistant acid phosphatase, matrix metalloproteinase-2 and tissue inhibitor of metalloproteinase-2 in early stages of canine osteoarthritis

Abstract: The aim of this study was to determine if the activity of tartrate-resistant acid phosphatase (TRAP) in the synovial fluid (SF) and serum can be used as a marker for diagnosing the early stages of osteoarthritis (OA). We also wished to determine if identifiable differences in the concentrations of matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) could be detected in SF between normal joints and OA joints for the diagnosis of early OA. Ten skeletally mature beagle dogs underwent a unilateral surgical transection of the cranial cruciate ligament and medial meniscectomy. Five sham-operated beagle dogs were used as controls. The synovial fluid was collected in 1, 2 and 3 months and examined by western blotting for MMP-2 and ELISA for TIMP-2. The activity of TRAP in the SF and serum was measured using a spectrophotometer. In addition, the presence of TRAP positive cells in the synovium was identified by enzyme histochemistry. The level of the activity of TRAP and MMP-2 in the SF from the induced OA dogs was significantly higher than that of the control over a three-month period ( P < 0.05). The TIMP-2 level in the SF was significantly lower in the induced OA dogs than in the control. However, there was no difference in TRAP activity in the serum. Histochemistry revealed a higher number of TRAP positive cells in the synovium from the induced OA dogs. Based on these data, we conclude that the activity of TRAP, MMP-2 and TIMP-2 in SF can be used as a biomarker to diagnose and monitor the early stages of OA.

Inhibition of RANKL-stimulated osteoclast differentiation by Schisandra chinensis through down-regulation of NFATc1 and c-fos expression.

Abstract: Schisandra chinenesis (SC) has been reported to have ameliorative effect on osteoporosis. However, the mechanisms underlying the anti-osteoporosis activity of SC have not been clearly elucidated. In the present study, we determined the effects of SC on The receptor activator of NF-kB ligand (RANKL)-induced osteoclastogenesis and its potential mechanism. Raw 264.7 cells were treated with 0.6, 6 and 60 μg/mL SC in the presence of 100 ng/mL RANKL for 7 days. RANKL-induced osteoclast formation was analyzed by tartrate resistant acid phosphatase (TRAP) staining. The osteoclast differentiation-related factors were confirmed along with TNF-α. SC inhibits the RANKL-induced osteoclast differentiation in dose-dependent manner within non-toxic concentrations. The supernatant concentrations of TNF-α were significantly decreased by SC treatment. In addition, osteoclastogenesis-related factors, TRAP6 and NF-κB, were markedly decreased by SC in RANKL-induced osteoclasts. Mechanistically, SC reduced the RANKL-triggered NFATc1 and c-fos expressions. Taken together, our data suggest that SC can modulate bone metabolism by suppressing RANKL-induced osteoclast differentiation.

Treatment of collagenase-induced osteoarthritis with a viral vector encoding TSG-6 results in ectopic bone formation.

Abstract: Objective Tumor necrosis factor-inducible gene 6 (TSG-6) has anti-inflammatory and chondroprotective effects in mouse models of inflammatory arthritis. Because cartilage damage and inflammation are also observed in osteoarthritis (OA), we determined the effect of viral overexpression of TSG-6 in experimental osteoarthritis. Methods Bone marrow-derived cells were differentiated to multinucleated osteoclasts in the presence of recombinant TSG-6 or after transduction with a lentiviral TSG-6 expression vector. Multi-nucleated osteoclasts were analyzed after tartrate resistant acid phosphatase staining and resorption activity was determined on dentin slices. Collagenase-induced osteoarthritis (CIOA) was induced in C57BL/6 mice after intra-articular injection of an adenoviral TSG-6 or control luciferase expression vector. Inflammation-related protease activity was measured using bioluminescent Prosense probes. After a second adenovirus injection, cartilage damage was assessed in histological sections stained with Safranin-O. Ectopic bone formation was scored in X-ray images of the affected knees. Results TSG-6 did not inhibit the formation of multi-nucleated osteoclasts, but caused a significant reduction in the resorption activity on dentin slices. Adenoviral TSG-6 gene therapy in CIOA could not reduce the cartilage damage compared to the luciferase control virus and no significant difference in inflammation-related protease activity was noted between the TSG-6 and control treated group. Instead, X-ray analysis and histological analysis revealed the presence of ectopic bone formation in the TSG-6 treated group. Conclusion Gene therapy based on the expression of TSG-6 could not provide cartilage protection in experimental osteoarthritis, but instead resulted in increased ectopic bone formation.

Vitamin C reverses bone loss in an osteopenic rat model of osteoporosis

Abstract: Fruits and vegetables are rich in vitamin C with antioxidant properties which are known to influence bone quality. This study evaluated whether vitamin C (1000 mg/L) added to drinking water reverses the bone loss in ovariectomized rats. Ninety-day-old female Sprague-Dawley rats were randomly assigned to either sham (n = 14) or ovariecotmized groups (n = 28). Sixty days after ovariectomy, the treatments were sham, ovariectomy (OVX), OVX + vitamin C (22 mg oral intake daily) for 60 days. Urine was collected for deoxypyridinoline (DPD) evaluation, rats were sacrificed, and antioxidant capacity, osteopontin, alkaline phosphatase (ALP), and bone specific tartrate resistant acid phosphatase (TRAP) were evaluated in the plasma. Right femur and 5th lumbar were evaluated for bone density, strength, ash, Ca, and Mg concentrations. Antioxidant capacity, ALP activity, osteopontin decreased (p-value 0.1) femoral strength, ash, or Ca, and Mg concentrations, while it increased (p-value < 0.05) the 5th lumbar density, ash, and Ca and Mg concentrations. In conclusion, vitamin C increased bone quality and antioxidant capacity in ovariectomized rats.

Abnormal bone remodelling activity of dental follicle cells from a cleidocranial dysplasia patient.

Abstract: Objectives To explore the role of dental follicle cells (DFCs) with a novel cleidocranial dysplasia (CCD) causative gene RUNX2 mutation (DFCsRUNX2+/m ) in delayed permanent tooth eruption. Materials and methods A CCD patient with typical clinical features was involved in this study. DFCsRUNX2+/m were cultured and DNA was extracted for RUNX2 mutation screening. Measurements of cell proliferation, alkaline phosphatase (ALP) activity, alizarin red staining and osteoblast-specific genes expression were performed to assess osteogenesis of DFCsRUNX2+/m . Co-culture of DFCs and peripheral blood mononuclear cells (PBMCs), followed tartrate-resistant acid phosphatase (TRAP) staining, real-time PCR and western blot were performed to evaluate osteoclast-inductive capacity of DFCsRUNX2+/m . Results A missense RUNX2 mutation (c. 557G>C) was found in DFCsRUNX2+/m from the CCD patient. Compared with normal controls, this mutation did not affect the proliferation of DFCsRUNX2+/m , but down-regulated the expression of osteogenesis-related genes, leading to a decrease in ALP activity and mineralisation. Co-culture results showed that DFCsRUNX2+/m reduced the formation of TRAP+ multinucleated cells and the expression of osteoclastogenesis-associated genes. Furthermore, the mutation reduced the ratio of RANKL/OPG in DFCsRUNX2+/m . Conclusions DFCsRUNX2+/m disturbs bone remodelling activity during tooth eruption through RANK/RANKL/OPG signalling pathway and may thus be responsible for impaired permanent tooth eruption in CCD patients.

Biological characteristics of osteoclast exosomes and their role in the osteogenic differentiation of somatic cells prior to osteogenesis.

Abstract: This study aimed to investigate the biological characteristics of osteoclast exocrine bodies and their role in the differentiation of somatic cells, so as to find out the key factors involved in osteoclast exosomatic growth and osteogenesis. RANKL (Receptor Activator for Nuclear Factor-κ B Ligand) induced factor was used to induce the osteoclast differentiation of Raw 264.7 cells, and TRAP (Tartrate resistant acid phosphatase) staining was employed to identify induced cells. Ultra-filtration centrifugation was used to separate OC-exosomes from osteoclast supernatant, while Western blot was employed to detect the expression characteristics of exosomal proteins CD9 and CD63. PKH67 labeled exosomes were observed to target kusao cells, which were divided into 3 groups, i.e., the complete medium group (group A), the osteoblast induced group (group B), and the osteogenesis induced liquid + OC-exosomes group (group C). The medium was changed on the next day and after 14-day culture. Using Western blot, alizarin red staining and Von Kossa silver staining, the role of OC-exosomes in the differentiation of kusao cells was clarified. Results showed that TRAP staining showed osteoclasts as irregular and TRAP positive giant cells with a red multicore and a large volume. Microcapsule membrane structures with a uniform size were detected in osteoclast supernatant, and the expression of CD9 and CD63 proteins was confirmed by Western blot. In addition, the Western blot results showed that the expression of RUNX2 (Runt-related transcription factor 2) protein in group B was 1.254 times of that in group A and 2.636 times of that in group C. Furthermore, alizarin red staining showed that the ratios of calcium salt deposition area to the total area in group A, group B and group C were 0.208%, 3.469%, and 20.724%, respectively. Von Kossa silver staining showed that the ratios of calcium salt deposition area to the total area in group A, group B and group C were 0.064%, 2.636%, and 20.872%, respectively. To sum up, OC-exosomes can promote the osteogenic differentiation of osteoblast cells (kusao cells).

Bone Degeneration, Inflammation and Secondary Complications of Arthritis: Potential Targets and their Natural Inhibitors.

Abstract: Arthritis is marked by joint deterioration that affects articular cartilage and subchondral bone. Though cartilage degradation does the major damage during arthritis, subsequent bone degeneration cannot be neglected. Recent progress in arthritis research has identified the clinical importance of bone erosion in destructive arthritis. Studies have showed the key role played by osteoclasts and receptor activator of nuclear factor kappaB ligand (RANKL) signaling in bone erosion. Cathepsins and tartrate resistant acid phosphatase (TRAP) are considered key enzymatic factors contributing to bone erosion. Further, reactive oxygen species (ROS) formed at the ruffled border of osteoclasts also causes bone resorption and matrix degradation. Besides, severe inflammation during arthritis induces bone erosion by aiding in Ca2+ removal and activating osteoclastogenesis. The inflammatory cytokines and ROS influence osteoclast differentiation by regulating osteoclast-lineage cells or by acting on other cells to regulate the expression of RANKL and osteoprotegerin (OPG). The enhanced production of pro-inflammatory cytokines and ROS in arthritis stimulates tissue injury by means of oxidative damage leading to vital organ damage and synovial and circulatory cell apoptosis. Thus, blocking enzymatic and non-enzymatic factors responsible for bone erosion and inflammation is considered a prime strategy in the management of arthritis. In this review we provide an overview of the mechanisms of bone erosion, inflammation and associated oxidative stress/damage during arthritis perpetuation along with shedding light on potential targets. The article also describes the possible natural therapeutic agents that could prevent bone loss and inflammation, and related secondary complications of arthritis.

Anemarrhena asphodeloides Bunge ameliorates osteoporosis by suppressing osteoclastogenesis

Abstract: Anemarrhena asphodeloides Bunge has been traditionally used in Korean medicine for its antipyretic, diuretic, sedative, and antitussive effects. In the present study, the effects of an ethanol extract of A. asphodeloides Bunge (AAB) on osteoporosis and its underlying mechanisms on bone remodeling were investigated. Osteoporosis was induced in ICR strain mice by ovariectomy. The mice were divided into four groups: sham, ovariectomized, 17β‑estradiol and 100 mg/kg AAB. The treatment was continued for 4 weeks. Bone mineral density (BMD) and bone mineral content (BMC) were measured using dual‑energy X‑ray absorptiometry. In addition, Raw 264.7 cells were treated in the presence of 0.1, 1 and 10 µg/ml AAB with 100 ng/ml receptor activator of nuclear factor κΒ ligand (RANKL) to induce osteoclast formation and stained with tartrate resistant acid phosphatase. In addition, levels of osteoclast‑related factors were analyzed to investigate the signaling cascades in osteoclasts. The results demonstrated that AAB treatment reversed the decreases of both BMD and BMC in osteoporotic femurs. Additionally, the formation of osteoclasts was significantly suppressed by the AAB treatment in RANKL‑stimulated Raw 264.7 cells. Compared with cells treated with RANKL alone, the AAB‑treated osteoclasts had significantly decreased tumor necrosis factor‑α and interleukin‑6. The protein levels of c‑fos were also decreased in the AAB‑treated osteoclasts. Furthermore, the RANKL‑induced nuclear translocation of nuclear factor‑κB was attenuated in osteoclasts by the AAB treatment compared with cells treated with RANKL alone. Finally, AAB treatment downregulated the phosphorylation of mitogen‑activated protein kinases. The present results demonstrated that AAB exhibited ameliorative effects on osteoporosis by inhibiting osteoclastogenesis, and suggested that AAB may be a potential candidate for the treatment of osteoporosis.

Effects of pressurized steam-treated Corni Fructus extract on osteoblast differentiation and osteoclast formation

Abstract: This study investigated the effects of pressurized steam-treated Corni Frutus (PSC) extract on osteoblast differentiation and osteoclast formation. The osteoblast differentiation effect of the extract was evaluated by measuring cellular alkaline phosphatase (ALP) activity, cell matrix ALP staining, alizarin Red S staining and von Kossa staining on proliferating MC3T3-E1 osteoblast cells. The results confirmed that ALP activity, cell matrix ALP staining, alizarin Red S staining and von Kossa staining were all increased as proliferation increased from 1 to 14 days, without cytotoxicity. The osteoclast formation effect of the PSC extract was evaluated by measuring the cellular tartrate-resistant acid phosphatase (TRAP) activity and cell matrix TRAP staining on receptor activator of nuclear factor kappa-B ligand (RANKL)-induced RAW264.7 osteoclast cells. Treating RAW264.7 cells with RANKL for 7 days increased matrix staining for TRAP and cellular TRAP activity. The PSC extract decreased these changes in a concentration-dependent manner. Therefore, PSC is expected to be a natural source for developing health functional foods and medicinal agents to prevent bone-related diseases, such as osteoporosis, by increasing osteoblast differentiation and reducing osteoclast activity.

Application of photopigment in preparation of medicines for treating bone loss diseases

Abstract: The invention discloses application of photopigment in preparation of medicines for treating bone loss diseases. Tartrate resistant acid phosphatase dyeing, immunofluorescence, RT-PCR (reverse transcription-polymerase chain reaction) and western blot detection are performed by an inventor, so as to evaluate the influence to the osteoclast differentiation, osteoclast absorption and osteoblast differentiation by photopigment. Proofed by results, the photopigment has the advantages that the photopigment can be used for inhibiting the expression of an osteoclast marker gene; the osteoclast formation and absorption induced by RANKL are inhibited by inhibiting the RANKL-induced NFAT activating and calcium signal conduction; interestingly, the osteoblast differentiation is also accelerated by thephotopigment, which is proved by the up-regulated expression of osteoblast marker genes ALP, Runx 2 and Collal; the photopigment can reduce the number of osteoclasts to inhibit the osteoclast function, so as to prevent the bone loss of the ovariectomized mouse; the photopigment may be used as a candidate medicine for treating the bone loss diseases, especially osteoporosis.